Urokinase plasminogen activator (uPA), its high affinity receptor (uPAR/CD87) and PA inhibitors (PAI's) are widely used biomarkers of malignant transformation, tumor growth, propensity to metastasize, and prognosis. uPA and its surface receptor (uPAR) are implicated in proliferation, migration and invasion of malignant cells, which is pivotal in the progression of tumor development and dissemination. uPA is composed of three domains: the growth factor-like domain (GFD), which binds uPAR, the kringle and a protease domain which converts plasminogen into plasmin and cleaves some other substrates. Disabling the uPA/uPAR system has become a rich target for anti-cancer therapies. However, existing anti-cancer therapeutic approaches that block uPA catalytic activity or prevent uPA-uPAR binding provide incomplete inhibition of tumor growth. The reason why these approaches have not been more effective in vivo could lie in their failure to inhibit other highly relevant yet unidentified uPA-driven pathway(s). Our studies indicate that scuPA is translocated to the nucleus of mammalian cells in vitro and in vivo in a kringle-dependent and uPAR-independent manner, and we have identified one critical intermediary in the process, nucleolin. In this application we propose to delineate the functions of uPA within the nucleus with the long-term goal of relating its nuclear translocation and uPA-mediated gene transcription to uPA-mediated tumor cell proliferation and survival using prostate cancer as a model. In Aim 1 we will explore the hypothesis that translocation of scuPA to the nucleus stimulates cell proliferation and inhibits apoptosis in a kringle-dependent manner. In Aim 2 we will explore the hypothesis that scuPA modulates expression of genes that regulate proliferation and/or apoptosis through this translocation pathway. In Aim 3 we will identify novel intracellular partners for uPA that mediate its nuclear transport or signalling using affinity isolation combined with proteomics approach, as well as a yeast two-hybrid system. Together these studies will elucidate a new facet of uPA biology and should provide an opportunity to develop sensitive and selective biomarkers that differentiate the subset of prostate cancer with a high propensity to metastasize from tumors with less aggressive behavior, and identify novel therapeutic targets to spur development of innovative therapeutic strategies to prevent untoward tumor cell proliferation and dissemination. PUBLIC HEALTH RELEVANCE: Although urokinase (uPA) and its receptor strongly contribute to the dissemination of prostate cancer, the efficacy of existing approaches to disable this system is incomplete, possibly because uPA acts through additional uncharacterized pathways. We recently reported that uPA translocates to the nuclei of tumor cells and regulates cell proliferation and survival. We propose to explore this previously unrecognized pathway to help identify novel therapeutic targets and spur development of agents that help prevent untoward tumor cell proliferation and tissue dissemination.